Proteins containing the double-stranded RNA binding motif (dsRBM) play key regulatory roles in both prokaryotic and eukaryotic organisms. This domain binds to structured RNAs and participates in protein-protein interactions. The RNA-dependent protein kinase (PKR) and nuclear factor 90 (NF90) each contain two dsRBMs. PKR is a cellular protein kinase that is induced by interferon and regulates protein synthesis and signal transduction pathways involved in viral infection and cellular growth control. Accordingly, PKR is tightly regulated and receives signals from a variety of sources. Several isoforms of NF90 exist in cells, including the C-terminal variant NF110, and members of this protein family have been implicated as both regulators and substrates of PKR. The NF90 family proteins are heavily phosphorylated during mitosis and have been associated with the regulation of gene expression at several levels including transcription, translation, and RNA turnover. NF90 purifies as a heterodimer with nuclear factor 45 (NF45), which can also modulate the functions of both PKR and NF90. The overall objectives of this application are to develop an understanding of the structure and function of proteins in the NF90 family, to elucidate the functional relationships between NF90/110, NF45 and PKR, and to explore the role(s) of NF90 family proteins in virus-infected and normal cells. To these ends, we will use current technologies including mass spectrometry and RNA interference, to examine the following: 1. The regulation of PKR by NF90/110 and NF45 in human cells and the yeast model system. 2. The role of NF90/110 phosphorylation by PKR and in mitosis. 3. Protein complexes formed by NF90 isoforms and the changes brought about by viral infection, interferon treatment, and cell cycle block. 4. The functions of NF90 isoforms during infection with adenovirus and HIV.